Excitatory amino acid-induced toxicity has been implicated in the nerve cell loss associated with several significant clinical conditions, including stroke, mechanical trauma to the nervous system, amyotrophic lateral sclerosis, and epilepsy. Strategies aimed at reducing this excitotoxicity have focused on blocking excitatory amino acid receptors, particularly the NMDA receptor and group I metabotropic glutamate receptors (mGluRs), activation of group II and III mGluRs, and reducing synaptic glutamate. This research program focuses on the latter two strategies to design, synthesize and test new compounds that may have therapeutic potential for the treatment of excitotoxicity. Central to this approach is N-acetylaspartylyglutamate (NAAG), the most prevalent and widely distributed peptide transmitter in the mammalian central nervous system. NAAG is a selective agonist at group II metabotropic glutamate receptors. Activation of these receptors has been shown to be neuroprotective in vitro and in vivo. NAAG and related group II agonists inhibit transmitter release by a presynaptic mechanism. Equally important, NAAG is inactivated by peptidase activity in the synaptic space to produce glutamate and N-acetylaspartate The program will extend preliminary work in which NAAG analogues with a central urea group have been synthesized and tested for their ability to inhibit enzymes that hydrolyze NAAG, particularly glutamate carboxypeptidase II (GCPII). An aim is to identify novel NAAG-peptidase inhibitors that will increase the peptide concentration following synaptic release, thus increasing activation of group II receptors, while decreasing the release of glutamate from NAAG. In our current SAR work, we have identified several urea-based structures that exhibit nM potency in the inhibition of this peptidase. Based upon these lead compounds, and using rational drug design, we will prepare a library of compounds that serve to further delineate structural features relevant to enhancing inhibitory potency and bioavailability. The program aims to acquire a better understanding of the typography of the enzyme active site of GCPII, and ultimately to produce compounds for use in vivo. Homology-based modeling methods will be employed to create a 3D structure of the enzyme, and the model will be used in turn to assist in ligand design. Since this drug design program has used the peptide as lead compound, it is anticipated that some of the peptidase inhibitors that are identified also may function as group II receptor agonists, an activity that would enhance their neuroprotective potential.